Ehf controls mammary alveolar lineage differentiation and is a putative suppressor of breast tumorigenesis.

The transcription factor EHF is highly expressed in the lactating mammary gland, but its role in mammary development and tumorigenesis is not fully understood. Utilizing a mouse model of Ehf deletion, herein, we demonstrate that loss of Ehf impairs mammary lobuloalveolar differentiation at late pregnancy, indicated by significantly reduced levels of milk genes and milk lipids, fewer differentiated alveolar cells, and an accumulation of alveolar progenitor cells. Further, deletion of Ehf increased proliferative capacity and attenuated prolactin-induced alveolar differentiation in mammary organoids. Ehf deletion also increased tumor incidence in the MMTV-PyMT mammary tumor model and increased the proliferative capacity of mammary tumor organoids, while low EHF expression was associated with higher tumor grade and poorer outcome in luminal A and basal human breast cancers. Collectively, these findings establish EHF as a non-redundant regulator of mammary alveolar differentiation and a putative suppressor of mammary tumorigenesis.

BECLIN1 is essential for intestinal homeostasis involving autophagy-independent mechanisms through its function in endocytic trafficking.

Autophagy-related genes have been closely associated with intestinal homeostasis. BECLIN1 is a component of Class III phosphatidylinositol 3-kinase complexes that orchestrate autophagy initiation and endocytic trafficking. Here we show intestinal epithelium-specific BECLIN1 deletion in adult mice leads to rapid fatal enteritis with compromised gut barrier integrity, highlighting its intrinsic critical role in gut maintenance. BECLIN1-deficient intestinal epithelial cells exhibit extensive apoptosis, impaired autophagy, and stressed endoplasmic reticulum and mitochondria. Remaining absorptive enterocytes and secretory cells display morphological abnormalities. Deletion of the autophagy regulator, ATG7, fails to elicit similar effects, suggesting additional novel autophagy-independent functions of BECLIN1 distinct from ATG7. Indeed, organoids derived from BECLIN1 KO mice show E-CADHERIN mislocalisation associated with abnormalities in the endocytic trafficking pathway. This provides a mechanism linking endocytic trafficking mediated by BECLIN1 and loss of intestinal barrier integrity. Our findings establish an indispensable role of BECLIN1 in maintaining mammalian intestinal homeostasis and uncover its involvement in endocytic trafficking in this process. Hence, this study has important implications for our understanding of intestinal pathophysiology.

Intestinal-specific Hdac3 deletion increases susceptibility to colitis and small intestinal tumor development in mice fed a high-fat diet.

High-fat (HF) diets (HFDs) and inflammation are risk factors for colon cancer; however, the underlying mechanisms remain to be fully elucidated. The transcriptional corepressor HDAC3 has recently emerged as a key regulator of intestinal epithelial responses to diet and inflammation with intestinal-specific Hdac3 deletion (Hdac3IKO) in mice increasing fatty acid oxidation genes and the rate of fatty acid oxidation in enterocytes. Hdac3IKO mice are also predisposed to experimentally induced colitis; however, whether this is driven by the intestinal metabolic reprogramming and whether this predisposes these mice to intestinal tumorigenesis is unknown. Herein, we examined the effects of intestinal-specific Hdac3 deletion on colitis-associated intestinal tumorigenesis in mice fed a standard (STD) or HFD. Hdac3IKO mice were highly prone to experimentally induced colitis, which was further enhanced by an HFD. Hdac3 deletion also accelerated intestinal tumor development, specifically when fed an HFD and most notably in the small intestine where lipid absorption is maximal. Expression of proteins involved in fatty acid metabolism and oxidation (SCD1, EHHADH) were elevated in the small intestine of Hdac3IKO mice fed an HFD, and these mice displayed increased levels of lipid peroxidation, DNA damage, and apoptosis in their villi, as well as extensive expansion of the stem cell and progenitor cell compartment. These findings reveal a novel role for Hdac3 in suppressing colitis and intestinal tumorigenesis, particularly in the context of consumption of an HFD, and reveal a potential mechanism by which HFDs may increase intestinal tumorigenesis by increasing fatty acid oxidation, DNA damage, and intestinal epithelial cell turnover.NEW & NOTEWORTHY We reveal a novel role for the transcriptional corepressor Hdac3 in suppressing colitis and intestinal tumorigenesis, particularly in the context of consumption of an HFD, and reveal a potential mechanism by which HFDs may increase intestinal tumorigenesis by increasing fatty acid oxidation, DNA damage, and intestinal epithelial cell turnover. We also identify a unique mouse model for investigating the complex interplay between diet, metabolic reprogramming, and tumor predisposition in the intestinal epithelium.

Genotype-Tailored ERK/MAPK Pathway and HDAC Inhibition Rewires the Apoptotic Rheostat to Trigger Colorectal Cancer Cell Death.

The EGFR/RAS/MEK/ERK signaling pathway (ERK/MAPK) is hyperactivated in most colorectal cancers. A current limitation of inhibitors of this pathway is that they primarily induce cytostatic effects in colorectal cancer cells. Nevertheless, these drugs do induce expression of proapoptotic factors, suggesting they may prime colorectal cancer cells to undergo apoptosis. As histone deacetylase inhibitors (HDACis) induce expression of multiple proapoptotic proteins, we examined whether they could synergize with ERK/MAPK inhibitors to trigger colorectal cancer cell apoptosis. Combined MEK/ERK and HDAC inhibition synergistically induced apoptosis in colorectal cancer cell lines and patient-derived tumor organoids in vitro, and attenuated Apc-initiated adenoma formation in vivo. Mechanistically, combined MAPK/HDAC inhibition enhanced expression of the BH3-only proapoptotic proteins BIM and BMF, and their knockdown significantly attenuated MAPK/HDAC inhibitor-induced apoptosis. Importantly, we demonstrate that the paradigm of combined MAPK/HDAC inhibitor treatment to induce apoptosis can be tailored to specific MAPK genotypes in colorectal cancers, by combining an HDAC inhibitor with either an EGFR, KRASG12C or BRAFV600 inhibitor in KRAS/BRAFWT; KRASG12C, BRAFV600E colorectal cancer cell lines, respectively. These findings identify a series of ERK/MAPK genotype-tailored treatment strategies that can readily undergo clinical testing for the treatment of colorectal cancer.

Epithelial de-differentiation triggered by co-ordinate epigenetic inactivation of the EHF and CDX1 transcription factors drives colorectal cancer progression.

Colorectal cancers (CRCs) often display histological features indicative of aberrant differentiation but the molecular underpinnings of this trait and whether it directly drives disease progression is unclear. Here, we identify co-ordinate epigenetic inactivation of two epithelial-specific transcription factors, EHF and CDX1, as a mechanism driving differentiation loss in CRCs. Re-expression of EHF and CDX1 in poorly-differentiated CRC cells induced extensive chromatin remodelling, transcriptional re-programming, and differentiation along the enterocytic lineage, leading to reduced growth and metastasis. Strikingly, EHF and CDX1 were also able to reprogramme non-colonic epithelial cells to express colonic differentiation markers. By contrast, inactivation of EHF and CDX1 in well-differentiated CRC cells triggered tumour de-differentiation. Mechanistically, we demonstrate that EHF physically interacts with CDX1 via its PNT domain, and that these transcription factors co-operatively drive transcription of the colonic differentiation marker, VIL1. Compound genetic deletion of Ehf and Cdx1 in the mouse colon disrupted normal colonic differentiation and significantly enhanced colorectal tumour progression. These findings thus reveal a novel mechanism driving epithelial de-differentiation and tumour progression in CRC.

Changing lung function and associated health-related quality-of-life: A five-year cohort study of Malawian adults.

BACKGROUND: In Sub-Saharan Africa cross-sectional studies report a high prevalence of abnormal lung function indicative of chronic respiratory disease. The natural history and health impact of this abnormal lung function in low-and middle-income countries is largely unknown. METHODS: A cohort of 1481 adults representative of rural Chikwawa in Malawi were recruited in 2014 and followed-up in 2019. Respiratory symptoms and health-related quality of life (HRQoL) were quantified. Lung function was measured by spirometry. FINDINGS: 1232 (83%) adults participated; spirometry was available for 1082 (73%). Mean (SD) age 49.5 (17.0) years, 278(23%) had ever smoked, and 724 (59%) were women. Forced expiratory volume in one second (FEV1) declined by 53.4 ml/year (95% CI: 49.0, 57.8) and forced vital capacity (FVC) by 45.2 ml/year (95% CI: 39.2, 50.5) . Chronic airflow obstruction increased from 9.5% (7.6, 11.6%) in 2014 to 17.5% (15.3, 19.9%) in 2019. There was no change in diagnosed asthma or in spirometry consistent with asthma or restriction. Rate of FEV1 decline was not associated with diagnosed Chronic obstructive pulmonary disease (COPD), asthma, or spirometry consistent with asthma, COPD, or restriction. HRQoL was adversely associated with respiratory symptoms (dyspnoea, wheeze, cough), previous tuberculosis, declining FEV1 and spirometry consistent with asthma or restriction. These differences exceeded the minimally important difference. INTERPRETATION: In this cohort, the increasing prevalence of COPD is associated with the high rate of FEV1 decline and lung function deficits present before recruitment. Respiratory symptoms and sub-optimal lung function are independently associated with reduced HRQoL.

EHF is essential for epidermal and colonic epithelial homeostasis, and suppresses Apc-initiated colonic tumorigenesis.

Ets homologous factor (EHF) is a member of the epithelial-specific Ets (ESE) family of transcription factors. To investigate its role in development and epithelial homeostasis, we generated a series of novel mouse strains in which the Ets DNA-binding domain of Ehf was deleted in all tissues (Ehf-/-) or specifically in the gut epithelium. Ehf-/- mice were born at the expected Mendelian ratio, but showed reduced body weight gain, and developed a series of pathologies requiring most Ehf-/- mice to reach an ethical endpoint before reaching 1 year of age. These included papillomas in the facial skin, abscesses in the preputial glands (males) or vulvae (females), and corneal ulcers. Ehf-/-mice also displayed increased susceptibility to experimentally induced colitis, which was confirmed in intestinal-specific Ehf knockout mice. Gut-specific Ehf deletion also impaired goblet cell differentiation, induced extensive transcriptional reprogramming in the colonic epithelium and enhanced Apc-initiated adenoma development. The Ets DNA-binding domain of EHF is therefore essential for postnatal homeostasis of the epidermis and colonic epithelium, and its loss promotes colonic tumour development.

Cohort profile: The Chikwawa lung health cohort; a population-based observational non-communicable respiratory disease study of adults in Malawi.

PURPOSE: The aim of this article is to provide a detailed description of the Chikwawa lung health cohort which was established in rural Malawi to prospectively determine the prevalence and causes of lung disease amongst the general population of adults living in a low-income rural setting in Sub-Saharan Africa. PARTICIPANTS: A total of 1481 participants were randomly identified and recruited in 2014 for the baseline study. We collected data on demographic, socio-economic status, respiratory symptoms and potentially relevant exposures such as smoking, household fuels, environmental exposures, occupational history/exposures, dietary intake, healthcare utilization, cost (medication, outpatient visits and inpatient admissions) and productivity losses. Spirometry was performed to assess lung function. At baseline, 56.9% of the participants were female, mean age was 43.8 (SD:17.8) and mean body mass index (BMI) was 21.6 Kg/m2 (SD: 3.46). FINDINGS TO DATE: The cohort has reported the prevalence of chronic respiratory symptoms (13.6%, 95% confidence interval [CI], 11.9-15.4), spirometric obstruction (8.7%, 95% CI, 7.0-10.7), and spirometric restriction (34.8%, 95% CI, 31.7-38.0). Additionally, an annual decline in forced expiratory volume in one second [FEV1] of 30.9mL/year (95% CI: 21.6 to 40.1) and forced vital capacity [FVC] by 38.3 mL/year (95% CI: 28.5 to 48.1) has been reported. FUTURE PLANS: The ongoing phases of follow-up will determine the annual rate of decline in lung function as measured through spirometry and the development of airflow obstruction and restriction, and relate these to morbidity, mortality and economic cost of airflow obstruction and restriction. Population-based mathematical models will be developed driven by the empirical data from the cohort and national population data for Malawi to assess the effects of interventions and programmes to address the lung burden in Malawi. The present follow-up study started in 2019.

Exploring perspectives on chronic obstructive pulmonary disease in people who smoke heroin: a qualitative study.

BACKGROUND: Smoking rather than injecting heroin has become more common over the last 20 years. Although there is an increasing body of evidence describing high levels of chronic obstructive pulmonary disease (COPD) in people who smoke heroin, there is limited evidence documenting the impact of the long-term condition on this population group. AIM: This study aimed to describe the experiences of people who smoke heroin with COPD in Liverpool, UK. DESIGN & SETTING: Participants were purposefully sampled for this qualitative study. They included adults enrolled in an opioid replacement clinic run by Addaction in Liverpool, who had already engaged with spirometry testing for COPD as part of a previous study. METHOD: Semi-structured interviews were performed with participants with spirometrically confirmed COPD in opioid replacement clinics. Data were analysed using a framework analysis approach. RESULTS: Sixteen potential participants were invited to take part in the study, of which 10 agreed and were interviewed. Three themes common to all interviews were identified: functional measures of lung health that impacted on their activities of daily living; inhaler and medication perceptions with erratic use that was not concordant with their prescription; and the impact of difficulties accessing care. CONCLUSION: These findings, along with previous studies highlighting the prevalence of COPD in this population, warrant efforts to integrate community COPD and opioid replacement services to improve outcomes for this vulnerable population.

Non-communicable respiratory disease and air pollution exposure in Malawi: a prospective cohort study.

RATIONALE: There are no population-based studies from sub-Saharan Africa describing longitudinal lung function in adults. OBJECTIVES: To explore the lung function trajectories and their determinants, including the effects of air pollution exposures and the cleaner-burning biomass-fuelled cookstove intervention of the Cooking and Pneumonia Study (CAPS), in adults living in rural Malawi. METHODS: We assessed respiratory symptoms and exposures, spirometry and measured 48-hour personal exposure to fine particulate matter (PM2.5) and carbon monoxide (CO), on three occasions over 3 years. Longitudinal data were analysed using mixed-effects modelling by maximum likelihood estimation. MEASUREMENTS AND MAIN RESULTS: We recruited 1481 adults, mean (SD) age 43.8 (17.8) years, including 523 participants from CAPS households (271 intervention; 252 controls), and collected multiple spirometry and air pollution measurements for 654 (44%) and 929 (63%), respectively. Compared with Global Lung Function Initiative African-American reference ranges, mean (SD) FEV1 (forced expiratory volume in 1 s) and FVC (forced vital capacity) z-scores were -0.38 (1.14) and -0.19 (1.09). FEV1 and FVC were determined by age, sex, height, previous TB and body mass index, with FEV1 declining by 30.9 mL/year (95% CI: 21.6 to 40.1) and FVC by 38.3 mL/year (95% CI: 28.5 to 48.1). There was decreased exposure to PM2.5 in those with access to a cookstove but no effect on lung function. CONCLUSIONS: We did not observe accelerated lung function decline in this cohort of Malawian adults, compared with that reported in healthy, non-smoking populations from high-income countries; this suggests that the lung function deficits we measured in adulthood may have origins in early life.

Overexpression of TP53 protein is associated with the lack of adjuvant chemotherapy benefit in patients with stage III colorectal cancer.

TP53 mutations drive colorectal cancer development, with missense mutations frequently leading to accumulation of abnormal TP53 protein. TP53 alterations have been associated with poor prognosis and chemotherapy resistance, but data remain controversial. Here, we examined the predictive utility of TP53 overexpression in the context of current adjuvant treatment practice for patients with stage III colorectal cancer. A prospective cohort of 264 stage III patients was tested for association of TP53 expression with 5-year disease-free survival, grouped by adjuvant treatment. Findings were validated in an independent retrospective cohort of 274 stage III patients. Overexpression of TP53 protein (TP53+) was found in 53% and 52% of cases from the prospective and retrospective cohorts, respectively. Among patients receiving adjuvant chemotherapy, TP53+ status was associated with shorter disease-free survival (p ≤ 0.026 for both cohorts), while no difference in outcomes between TP53+ and TP53- cases was observed for patients treated with surgery alone. Considering patients with TP53- tumors, those receiving adjuvant treatment had better outcomes compared with those treated with surgery alone (p ≤ 0.018 for both cohorts), while no treatment benefit was apparent for patients with TP53+ tumors. Combined cohort-stratified analysis adjusted for clinicopathological variables and DNA mismatch repair status confirmed a significant interaction between TP53 expression and adjuvant treatment for disease-free survival (pinteraction = 0.030). For the combined cohort, the multivariate hazard ratio for TP53 overexpression among patients receiving adjuvant chemotherapy was 2.03 (95% confidence interval 1.41-2.95, p < 0.001), while the hazard ratio for adjuvant treatment among patients with TP53- tumors was 0.42 (95% confidence interval 0.24-0.71, p = 0.001). Findings were maintained irrespective of tumor location or when restricted to mismatch repair-proficient tumors. Our data suggest that adjuvant chemotherapy benefit in stage III colorectal cancer is restricted to cases with low-level TP53 protein expression. Identifying TP53+ tumors could highlight patients that may benefit from more aggressive treatment or follow-up.

Screening Heroin Smokers Attending Community Drug Clinics for Change in Lung Function: A Cohort Study.

BACKGROUND: Heroin smokers have high rates of COPD, respiratory morbidity, hospital admission, and mortality. We assessed the natural history of symptoms and lung function in this population over time. METHODS: A cohort of heroin smokers with COPD was followed for 18 to 24 months. At baseline and follow-up, respiratory symptoms were measured by the Medical Research Council Dyspnea Scale (MRC) and the COPD Assessment Tool (CAT), and postbronchodilator spirometry was performed. Frequency of health-care-seeking episodes was extracted from routine health records. Parametric, nonparametric, and linear regression models were used to analyze the change in symptoms and lung function over time. RESULTS: Of 372 participants originally recruited, 161 were assessed at follow-up (mean age, 51.0 ± 5.3 years; 74 women [46%]) and 106 participants completed postbronchodilator spirometry. All participants were current or previous heroin smokers, and 122 (75.8%) had smoked crack. Symptoms increased over time (MRC score increased by 0.48 points per year, P < .001; CAT score increased by 1.60 points per year, P < .001). FEV1 declined annually by 90 ± 190 mL (P < .001). This deterioration was not associated with change in tobacco or heroin smoking status or use of inhaled medications. CONCLUSIONS: Heroin smokers experience a high and increasing burden of chronic respiratory symptoms and a decline in FEV1 that exceeds the normal age-related decline observed among tobacco smokers with COPD and healthy nonsmokers. Targeted COPD diagnostic and treatment services hosted within opiate substitution services could benefit this vulnerable, relatively inaccessible, and underserved group of people.

Noncommunicable Respiratory Disease and Air Pollution Exposure in Malawi (CAPS). A Cross-Sectional Study.

RATIONALE: Noncommunicable respiratory diseases and exposure to air pollution are thought to be important contributors to morbidity and mortality in sub-Saharan African adults. OBJECTIVES: We set out to explore the prevalence and determinants of noncommunicable respiratory disease among adults living in Chikhwawa District, Malawi. METHODS: We performed a cross-sectional study among adults in communities participating in a randomized controlled trial of a cleaner-burning biomass-fueled cookstove intervention (CAPS [Cooking and Pneumonia Study]) in rural Malawi. We assessed chronic respiratory symptoms, spirometric abnormalities, and personal exposure to air pollution (particulate matter <2.5 µm in aerodynamic diameter [PM2.5] and carbon monoxide [CO]). Weighted prevalence estimates were calculated; multivariable and intention-to-treat analyses were done. MEASUREMENTS AND MAIN RESULTS: One thousand four hundred eighty-one participants (mean [SD] age, 43.8 [17.8] yr; 57% female) were recruited. The prevalence of chronic respiratory symptoms, spirometric obstruction, and restriction were 13.6% (95% confidence interval [CI], 11.9-15.4), 8.7% (95% CI, 7.0-10.7), and 34.8% (95% CI, 31.7-38.0), respectively. Median 48-hour personal PM2.5 and CO exposures were 71.0 µg/m3 (interquartile range [IQR], 44.6-119.2) and 1.23 ppm (IQR, 0.79-1.93), respectively. Chronic respiratory symptoms were associated with current/ex-smoking (odds ratio [OR], 1.59; 95% CI, 1.05-2.39), previous tuberculosis (OR, 2.50; 95% CI, 1.04-15.58), and CO exposure (OR, 1.46; 95% CI, 1.04-2.05). Exposure to PM2.5 was not associated with any demographic, clinical, or spirometric characteristics. There was no effect of the CAPS intervention on any of the secondary trial outcomes. CONCLUSIONS: The burden of chronic respiratory symptoms, abnormal spirometry, and air pollution exposures in adults in rural Malawi is of considerable potential public health importance. We found little evidence that air pollution exposures were associated with chronic respiratory symptoms or spirometric abnormalities and no evidence that the CAPS intervention had effects on the secondary trial outcomes. More effective prevention and control strategies for noncommunicable respiratory disease in sub-Saharan Africa are needed. Clinical trial registered with www.isrctn.com (ISRCTN 59448623).

DUSP5 is methylated in CIMP-high colorectal cancer but is not a major regulator of intestinal cell proliferation and tumorigenesis.

The ERK signalling pathway regulates key cell fate decisions in the intestinal epithelium and is frequently dysregulated in colorectal cancers (CRCs). Variations in the dynamics of ERK activation can induce different biological outcomes and are regulated by multiple mechanisms, including activation of negative feedback loops involving transcriptional induction of dual-specificity phosphatases (DUSPs). We have found that the nuclear ERK-selective phosphatase DUSP5 is downregulated in colorectal tumours and cell lines, as previously observed in gastric and prostate cancer. The DUSP5 promoter is methylated in a subset of CRC cell lines and primary tumours, particularly those with a CpG island methylator phenotype (CIMP). However, this epigenetic change alone could not account for reduced DUSP5 expression in CRC cells. Functionally, DUSP5 depletion failed to alter ERK signalling or proliferation in CRC cell lines, and its transgenic overexpression in the mouse intestine had minimal impact on normal intestinal homeostasis or tumour development. Our results suggest that DUSP5 plays a limited role in regulating ERK signalling associated with the growth of colorectal tumours, but that methylation the DUSP5 gene promoter can serve as an additional means of identifying CIMP-high colorectal cancers.

ATF3 Repression of BCL-XL Determines Apoptotic Sensitivity to HDAC Inhibitors across Tumor Types.

Purpose: Histone deacetylase inhibitors (HDACi) are epigenome-targeting small molecules approved for the treatment of cutaneous T-cell lymphoma and multiple myeloma. They have also demonstrated clinical activity in acute myelogenous leukemia, non-small cell lung cancer, and estrogen receptor-positive breast cancer, and trials are underway assessing their activity in combination regimens including immunotherapy. However, there is currently no clear strategy to reliably predict HDACi sensitivity. In colon cancer cells, apoptotic sensitivity to HDACi is associated with transcriptional induction of multiple immediate-early (IE) genes. Here, we examined whether this transcriptional response predicts HDACi sensitivity across tumor type and investigated the mechanism by which it triggers apoptosis.Experimental Design: Fifty cancer cell lines from diverse tumor types were screened to establish the correlation between apoptotic sensitivity, induction of IE genes, and components of the intrinsic apoptotic pathway.Results: We show that sensitivity to HDACi across tumor types is predicted by induction of the IE genes FOS, JUN, and ATF3, but that only ATF3 is required for HDACi-induced apoptosis. We further demonstrate that the proapoptotic function of ATF3 is mediated through direct transcriptional repression of the prosurvival factor BCL-XL (BCL2L1) These findings provided the rationale for dual inhibition of HDAC and BCL-XL, which we show strongly cooperate to overcome inherent resistance to HDACi across diverse tumor cell types.Conclusions: These findings explain the heterogeneous responses of tumor cells to HDACi-induced apoptosis and suggest a framework for predicting response and expanding their therapeutic use in multiple cancer types. Clin Cancer Res; 23(18); 5573-84. ©2017 AACR.

Dual Targeting of Bromodomain and Extraterminal Domain Proteins, and WNT or MAPK Signaling, Inhibits c-MYC Expression and Proliferation of Colorectal Cancer Cells.

Inhibitors of the bromodomain and extraterminal domain (BET) protein family attenuate the proliferation of several tumor cell lines. These effects are mediated, at least in part, through repression of c-MYC. In colorectal cancer, overexpression of c-MYC due to hyperactive WNT/ß-catenin/TCF signaling is a key driver of tumor progression; however, effective strategies to target this oncogene remain elusive. Here, we investigated the effect of BET inhibitors (BETi) on colorectal cancer cell proliferation and c-MYC expression. Treatment of 20 colorectal cancer cell lines with the BETi JQ1 identified a subset of highly sensitive lines. JQ1 sensitivity was higher in cell lines with microsatellite instability but was not associated with the CpG island methylator phenotype, c-MYC expression or amplification status, BET protein expression, or mutation status of TP53, KRAS/BRAF, or PIK3CA/PTEN Conversely, JQ1 sensitivity correlated significantly with the magnitude of c-MYC mRNA and protein repression. JQ1-mediated c-MYC repression was not due to generalized attenuation of ß-catenin/TCF-mediated transcription, as JQ1 had minimal effects on other ß-catenin/TCF target genes or ß-catenin/TCF reporter activity. BETi preferentially target super-enhancer-regulated genes, and a super-enhancer in c-MYC was recently identified in HCT116 cells to which BRD4 and effector transcription factors of the WNT/ß-catenin/TCF and MEK/ERK pathways are recruited. Combined targeting of c-MYC with JQ1 and inhibitors of these pathways additively repressed c-MYC and proliferation of HCT116 cells. These findings demonstrate that BETi downregulate c-MYC expression and inhibit colorectal cancer cell proliferation and identify strategies for enhancing the effects of BETi on c-MYC repression by combinatorial targeting the c-MYC super-enhancer. Mol Cancer Ther; 15(6); 1217-26. ©2016 AACR.